AC amplification gain in organic electrochemical transistors for impedance-based single cell sensors

Research on electrolyte-gated and organic electrochemical transistor (OECT) architectures is motivated by the prospect of a highly biocompatible interface capable of amplifying bioelectronic signals at the site of detection. Despite many demonstrations in these directions, a quantitative model for OECTs as impedance biosensors is still lacking. We overcome this issue by introducing a model experiment where we simulate the detection of a single cell by the impedance sensing of a dielectric microparticle. The highly reproducible experiment allows us to study the impact of transistor geometry and operation conditions on device sensitivity. With the data we rationalize a mathematical model that provides clear guidelines for the optimization of OECTs as single cell sensors, and we verify the quantitative predictions in an in-vitro experiment. In the optimized geometry, the OECT-based impedance sensor allows to record single cell adhesion and detachment transients, showing a maximum gain of 20.2±0.9 dB with respect to a single electrode-based impedance sensor.

During impedance sensing measurements, the OECTs were biased with the DC gate and drain voltages at which their transconductance gm,DC takes its maximum value (the working point). We plot in Supplementary Figure 2 the DC trans-characteristic and transconductance curves of a WxL= 100x100 µm sensor. The latter were obtained by direct differentiation of the former and show a transconductance peak which increases with the absolute value of the applied DC drain voltage. The working point was set in correspondence of one of these maxima, choosing a configuration with both a reasonably high transconductance and low potential operation to limit material degradation and power consumption.
Supplementary Figure 2: setting the OECT working point for impedance sensing. a) Transfer characteristic of an OECT sensor and b) DC transconductance curves indicating the working point set for impedance sensing.

Supplementary Figure 3
The presence of the dielectric microparticle in close contact to the sensor surface hinders the ionic flow from the electrolyte into the sensing channel and thus increases the electrolyte impedance. This is widely observed in our experiment and reproduces in first approximation the basic working principle of impedance-based cell sensors. A qualitative demonstration of this observation is furtherly provided in the reported experiment ( Supplementary Figure 3a), where we measured the source current amplitude while gradually increasing the microparticle-channel distance d starting from the contact position. The microparticle displacement produces larger effect on the sensor response when the AFM probe is retracted for short distances from the contact position. At the same time, both the cantilever and the AFM stage changes the geometry of the liquid electrolyte and reasonably modify the electrolyte resistance Rel. Anyway, given the large diameter of the dielectric microparticle (50 µm) and the small displacement of the z-stage during the sensing experiment (5 µm), we expect that only the bottom part of the dielectric microparticle has an active role in modifying the ionic current flow, simulating in first order approximation a biological cell which adheres to the sensor surface. This is highlighted in Supplementary Figure 3b, where we report the OECT current variation per 1 µm-step as a function of d. We observe that larger effects are produced when the AFM probe is retracted for short distances from the contact position, indicating that the microparticle hindrance has a primary role in blocking the ionic flow from the electrolyte to the sensor channel.
Supplementary Figure 3: Measuring the microparticle displacement. a) Current amplitude variation in an OECT sensor (biased at its working point) when the microparticle is lifted from the contact position. The modulation frequency applied to the gate electrode is 1.17 kHz. b) OECT current variation per 1 µm-step as a function of the microparticle-channel distance d.
We report in Supplementary Figure 4 the normalized microparticle distance-AC current measurement shown in absolute value in Figure 1f. The steepest line acquired with the OECT configuration results from its amplification gain, that increases the level of the measured signal. Naturally, also noise present in the signal will be amplified. Such intrinsic noise sources include thermal noise effects and shot noise effect due to the impedance of the measured system and noise effects due to the living activities of the cell. Accordingly, the OECT cannot improve on the signal to noise ratio when the noise level is only determined by these intrinsic factors. The role of gain in improving signal to noise ratio becomes important when noise is introduced by the data acquisition system. In our case such noise is minimized in both cases (microelectrode and OECT) due to the use of very sophisticated signal conditioning circuits. For this reason, both signal traces in Figure 1f have a comparable signal to noise ratio. We note that in a realistic application scenario, microelectrode impedance recordings would be deteriorated due to a limited digital resolution.

Supplementary Figure 4: normalized microparticle distance -AC current measurements.
Comparison between normalized microparticle distance -AC current measurements acquired in the OECT and microelectrode configurations. The absolute data are shown in Figure 1f.

Supplementary Figure 5
We report in Supplementary Figure 5a and 5b the impedance and the phase spectra of the PEDOT:PSS channels used in the experiments (acquired in the microelectrode configuration). Measurements were fitted with an equivalent RC circuit to extract the electrolyte resistance and the channel capacitance. This can be related to the device sensitivity as ∂I/∂Rel = ∂I/∂d×∂d/∂R. The coefficient p = ∂R/∂d is universal and does not change as a function of frequency or transistor or microelectrode parameters for each channel geometry. To determine its value, we measured the gate impedance ZG(d) in the microelectrode configuration at high modulation frequency (11.7 kHz). In such a condition, the impedance of the channel capacitance Zch=1/(iωCch) is negligible, and only the Rel(d) curve is consequently measured. This is confirmed by the phase of the acquired signals (see inset in Figure   S4b), which assumes values close to zero in that frequency regime. We report in Supplementary

Supplementary Table 1
We report in the following table the full set of parameters used to best fit the experimental data with the quantitative model for PEDOT:PSS-based impedance sensors. The channel capacitance Cch and the electrolyte resistance Rel were extracted by fitting the electrochemical impedance spectroscopies with a RC series circuit model. The OECT transconductance gm was calculated from the DC transfer characteristics.

Cch (nF)
Rel ( As expressed by Eq. 2, the OECT sensitivity is obtained by the sum of the microelectrode and the channel sensitivity, !" and #$ , respectively. The sensitivities are defined as the derivative of the AC current with respect to the cell induced changes in electrolyte impedance: . We first consider the AC current contribution generated in the OECT channel. OECTs working in AC operation exhibit low pass filtering properties, thereby the cutoff frequency fc can be calculated from !
, while I0 can be obtained from the low-frequency limit as I0 = gm*VG,AC.
In the microparticle sensing experiment, the electrolyte impedance Zel » Rel is a real number, allowing for a straightforward expression for the cutoff frequency: It is worth highlighting that fc corresponds to the frequency at which an OECT-based impedance sensor reaches its maximum sensitivity. This can be demonstrated by considering eq. 2 and setting stationary point conditions: The resulting expression for sOECT max is

Supplementary Figure 6
For the single cell detection experiment we microfabricated a linear array of 10 PEDOT:PSS channels with dimensions WxL=200x50 µm. In this way, we largely increased the probability that a single cell reached a single PEDOT:PSS channel by gravity after seeding. We report in Supplementary  During the single-cell detection experiment, the OECT-based impedance sensors showed a significative shift in the low-pass cutoff, which was recovered only after the treatment with trypsin.
To demonstrate that such effect was caused by the cell adhesion process and not by other effects in the experimental setup (sensor degradation/contamination of the cell culture medium), we acquired the current spectrum of a control device (with the same dimensions and operating parameters) placed in the same reservoir, but with no cell seeded on the sensing channel. Results are reported in Measurements are acquired in different electrolytes but confirm the correct behavior of the device after removal of the cell residuals.